Summary: After entry of HIV-1 nucleoprotein into the cytoplasm of a target cell, viral DNA is synthesized by the virally encoded viral reverse transcriptase, and forms a preintegration complex that is translocated to the host cell nucleus. The level of virus production is determined to a large extent by the efficiency of these early steps in viral replication. We have demonstrated that monoclonal antibody (mab) to an amino-acid transporter effectively prevents early steps of HIV-1 infection of T cells. Mab to the transporter does not affect cell surface CD4 or CXCR4 expression and does not block virus binding or early entry events. Antibody treated cells failed to translocate HIV-1 DNA to the nucleus following infection and exhibited a block in synthesis of late full length viral DNA products, but not early or first strand RT products. Antibody binding to the transporter disrupted the interaction between an intermediate cytoplasmic filament and viral DNA, but did not interfere with viral interactions with actin. Our results identify a novel relationship between amino-acid cell surface transporters and intermediate filaments in the regulation of reverse transcription and nuclear translocation of HIV-1 viral DNA. Strategies targeting amino-acid transporters on the cell surface may have potential for HIV-1 therapeutics and prevention.